One part of the INSITE project focused on developing a multi-omics workflow and a mechanistic model of the in vitro biology observed in the fluidic device. This included the development of single-cell sequencing analysis pipelines and the creation of a comprehensive bone development and homeostasis atlas based on scRNASeq datasets. An app accompanying this atlas has been developed for community use. Metabolomics studies on human Periosteum Derived Cells (hPDCs) identified several indicators for successful in vivo bone regeneration. An intracellular network model was established, resulting in two shareable outputs: a knowledge base in CellCollective and an executable chondrocyte model for external use. Additionally, tissue-level neotissue growth models were recalibrated for different substrates and integrated into a multiscale model.
A second part of the project centered on developing the INSITE fluidic device, resulting in a fully sensored prototype with inline pH, oxygen, and temperature sensors. Neotissue growth was studied in 3D printed calcium phosphate scaffolds, and a second microfluidics device was optimized to apply mechanical loading to cells in hydrogels. This device was used to examine the effects of mechanical loading on macrophages, chondrocytes, and osteoprogenitor cells, providing input for other parts of the work in INSITE.
A third part of the project involved analyzing early bone healing phases and emulating the biophysical context within the in vitro system. Computational models of vascularization and immune response were developed, including an agent-based inflammatory model, validated through dedicated experiments. Single-cell RNA sequencing experiments are currently being analyzed to quantify vascularization and early immune responses during fracture healing under various mechanical loading regimes.
Finally, puttiing together all the aformentioned elements, we developed spatio-temporally patterned constructs for bone defect healing, utilizing a high-end bioprinter to print 3D cell-laden constructs. Various hydrogel blends were developed to enhance printability and mechanical properties. In vivo experiments with these printed implants are ongoing. Biomaterials-based constructs were also optimized and are being prepared for translation to clinical use.
Overall, the technologies and tools developed have broad applications beyond the initial scope, including lymphangiogenesis studies in inflammatory conditions and multi-organ toxicity applications. This work has been presented at conferences and is in various stages of publication.
